In this specification we are concerned with emulsions comprising microdroplets of water in oil, generally surfactant-stabilised, used in a microfluidic device. One or more biological entities such as one or more living cells or particles may be incorporated into each droplet and then experiments performed within the droplet, for example to perform a biological assay. The technique is promising because the volume of a droplet is very small, typically in the order of picoliters, and thus the volume of biological material needed is small, and because microdroplets can be generated and processed, for example directly within the microfluidic device, at rates in excess of several thousands per second. Further, integrated active elements on the microfluidic device can be used to control individual droplets—for example we have previously described in WO2009/050512, technology which enables the extraction on-chip of the contents of microdroplets by incorporating them into a continuous stream. Other general background prior art on microdroplets can be found in patents/applications in the name of RainDance Technologies Inc., for example WO2008/063227.
Typically the oil composition comprises a fluorous and/or mineral oil and, preferably, a surfactant, for example at around 0.5-5% vol/vol. Use of a fluorous oil is particularly advantageous when the microdroplets contain living entities because fluorous oil is good at transporting oxygen to the microdroplets. The surfactant may be either polymeric or small molecule; for example surfactants derived from block co-polymers of perfluoroethers such as Krytox™ or polyethylene glycol (PEG) may be used.
General background prior art relating to emulsions for other applications (generally cosmetics) can be found in the following: U.S. Pat. Nos. 5,587,153; 6,017,546; WO2005/099661; US2004/081633; U.S. Pat. No. 6,379,682; US2002/172703. Further background prior art can be found in: US2005/087122; WO2004/038363; US2008/053205; and US2007/0275415.
The material or analyte within a microdroplet may comprise, for example, DNA, protein, peptide, beads, particles, crystals, micelles, macromolecules, material for an enzymatic assay, organelles, an organism such as cell for example a mammalian cell, yeast cell, algal cell or bacterium, a virus, a prion and so forth. Typically a droplet has a diameter in the range 1-120 μm although droplets may be larger (or smaller), giving a volume in the range nanoliters to femtoliters. In this specification we are particularly, but not exclusively concerned with microdroplets containing cells—the techniques we describe are particularly useful for cells but may, in principle, be applied to microdroplets containing other organisms/biological entities, beads or particles.
Hitherto microdroplet-based processing of cells has generally been limited to labelling the cells with a fluorescent marker activated under certain circumstances. Then, as a microdroplet passes through a channel of the microfluidic device the microdroplet is illuminated by a laser and fluorescence or the absence of fluorescence is detected and the microdroplet containing the cell is directed accordingly. The inventors have recognised, however, that it is possible to improve upon this technique.